The desirability of conducting high throughput analysis of very small samples, e.g., biological samples, has led to the development of microfluid chip devices. These devices, constructed using techniques such as photolithography, wet chemical etching, and thin film deposition, that are commonly used in the production of electronic chips, allow one to perform separation and analysis of samples as small as a few picoliters or less. Sophisticated microfluid chip devices enable precise mixing, separation, and reaction of samples in a integrated system. Generally, microfluid chip have a number of micrometer width channels connecting various reservoirs. Materials are manipulated through the channels and reservoirs using electrokinetic forces or other means.
Microfluid chips have been interfaced to electrospray ionization mass spectrometers (Xue et al., Anal. Chem. 69:426, 1997; Ramsey and Ramsey, Anal. Chem. 69:1174, 1997).
Electrospray ionization is used to produce ions for mass spectrometry analysis from large, complex molecules, for example, proteins and nucleic acid molecules. In electrospray ionization, a sample solution enters an electrospray chamber through a hollow needle which is maintained at a few kilovolts relative to the walls of the electrospray chamber. The electrical field charges the surface of the liquid emerging from the needle, dispersing it by Coulomb forces into a spray of fine, charged droplets. At this point the droplets become unstable and break into daughter droplets. This process is repeated as solvent continues to evaporate from each daughter droplet. Eventually, the droplets become small enough for the surface charge density to desorb ions from the droplets into the abient gas. These ions, which include cations or anions attached to solvent or solute species which are not themselves ions, are suitable for analysis by a mass spectrometer.
Xue et al. (supra) reported that a stable electrospray could be generated directly from a multichannel microfluid chip channel outlet port which opened at the flat edge of the microfluid chip. The voltage for electrospray ionization was applied from a buffer reservoir at the sample side with the mass spectrometer orifice grounded. Xue et al. reported that, because the electrospray plume was unstable when there was insufficient liquid flow at the channel outlet port, a syringe pump was required to provide adequate liquid flow.
Ramsey and Ramsey (supra) reported that a stable electrospray could be generated directly from a channel outlet port which ended in an opening at the flat edge of a microfluid chip using electroosmotic pumping. Ramsey and Ramsey obtained a mass spectrum from a 10 .mu.M solution of tetrabutylammonium iodide, reportedly consuming 30 fmol of sample.
The results reported by Xue et al. and Ramsey and Ramsey suggest that electrospray directly from the outlet port of a microfluid chip has relatively poor sensitivity compared to that required for the analysis of minute quantities of biological macromolecules.
The poor sensitivity of the Ramsey and Ramsey and Xue et al. flow cells is likely caused by a number of factors. First, because the outlet port is in a flat edge of the microfluid chip, ionization of droplets leaving the chip causes the spray to spread out prior to entering the injection port. Second, because the edge cut channel is very large, a stable electrospray does not form efficiently, resulting in inefficient ionization. In addition, the Xue et al. flow cell requires a pump to generate a driving force and the Ramsey and Ramsey flow cell requires a side-arm channel to generate a driving force. These features reduce sensitivity and make it difficult to conduct high throughput analysis of minute samples.